A large non-parasitic population of Savalia savaglia (Bertoloni, 1819) in the Boka Kotorska Bay (Montenegro).

The golden coral Savalia savaglia is a long-living ecosystem engineer of Mediterranean circalittoral assemblages, able to induce necrosis of gorgonians' and black corals' coenenchyme and grow on their cleaned organic skeleton. Despite its rarity, in Boka Kotorska Bay (Montenegro) a shallow population of more than 1000 colonies was recorded close to underwater freshwater springs, which create very peculiar environmental conditions. In this context, the species was extremely abundant at two sites, while gorgonians were rare. The abundance and size of S. savaglia colonies and the diversity of the entire benthic assemblage were investigated by photographic sampling in a depth range of 0-35 m. Several living fragments of S. savaglia spread on the sea floor and small settled colonies (< 5 cm high) suggested a high incidence of asexual reproduction and a non-parasitic behaviour of this population. This was confirmed by studying thin sections of the basal portion of the trunk where the central core, generally represented by the remains of the gorgonian host skeleton, was lacking. The S. savaglia population of Boka Kotorska Bay forms the unique Mediterranean assemblage of the species deserving the definition of animal forest. Recently, temporary mitigation measures for anthropogenic impact were issued by the Government of Montenegro. Nevertheless, due to the importance of the sites the establishment of a permanent Marine Protected Area is strongly recommended.

Evaluation of In Vitro Cytotoxic Potential of Avarol towards Human Cancer Cell Lines and In Vivo Antitumor Activity in Solid Tumor Models.

The goal of this study was to determine the activity in vitro and in vivo of avarol, a sesquiterpene hydroquinone originating from the Dysidea avara sponge from the south Adriatic Sea, against different cancer cell lines and two types of mouse carcinoma. To investigate the in vitro cytotoxicity, a human cervix adenocarcinoma cell line (HeLa), human colon adenocarcinoma (LS174), human non-small-cell lung carcinoma (A549), and a normal human fetal lung fibroblast cell line (MRC-5) were used. The in vivo antitumor activity was investigated against two transplantable mouse tumors, the Ehrlich carcinoma (EC) and cervical cancer (CC-5). The effect of avarol on cancer cell survival, which was determined by the microculture tetrazolium test, confirmed a significant in vitro potency of avarol against the investigated cell lines, without selectivity towards MRC-5. The highest cytotoxicity was exhibited against HeLa cancer cells (10.22 ± 0.28 µg/mL). Moreover, potent antitumor activity against two tumor models was determined, as the intraperitoneal administration of avarol at a dose of 50 mg/kg resulted in a significant inhibition of tumor growth in mice. After three administrations of avarol, a 29% inhibition of the EC growth was achieved, while in the case of CC-5, a 36% inhibition of the tumor growth was achieved after the second administration of avarol. Therefore, the results indicate that this marine sesquiterpenoid hydroquinone could be a promising bioactive compound in the development of new anticancer medicine.

Fucus spiralis extract and fractions: anticancer and pharmacological potentials.

PURPOSE: Sea macroalgae are an important source of biologically highly valuable compounds. The main aim of this study was to investigate the in vitro anticancer properties and chemical composition of the dichloromethane-methanol extract and three fractions of the Fucus spiralis from coastline of Morocco. METHODS: Fractions were made from dichloromethane:methanol (1:1) extract of Fucus spiralis: petroleum-ether, ethyl-acetate and n-butanol. Extract and fractions were screened for in vitro cytotoxicity by MTT assay against human cervical adenocarcinoma (HeLa), colorectal adenocarcinoma (LS-174T), lung carcinoma (A549), and normal human lung fibroblasts (MRC-5). Cell cycle distribution of the HeLa cells was evaluated using flow cytometry. Acridine orange (AO)-ethidium bromide (EB) staining was used to assess morphological changes of HeLa cells under fluorescence microscope. Anti-migration and anti-angiogenic properties were investigated using scratch and tube formation assays against human endothelium-derived permanent EA.hy926 cell line. Antidiabetic activity was tested using anti-α-glucosidase assay. Antimicrobial effect was tested using micro- dilution method. RESULTS: Petroleum-ether fraction оf Fucus spiralis rich in fatty acids exerted the highest cytotoxicity against HeLa cells. Ethyl-acetate and petroleum-ether fractions induced the highest accumulation of the HeLa cells in sub-G1 and G2/M phases. Extract and fractions showed proapoptotic effect on HeLa cells under fluorescent microscope. They exhibited antimigratory and antiangiogenic effects in vitro. IC50 value for α-glucosidase inhibitory activity was much stronger than standard acarbose. n-Butanol fraction exerted the highest antibacterial and antifungal activity. CONCLUSIONS: The investigation of various biological activities of the extract and fractions obtained from Fucus spiralis may suggest a promising anticancer and pharmacological potential of this edible macroalga.

Chemical Analysis of Selected Seaweeds and Seagrass from the Adriatic Coast of Montenegro.

Three seaweeds (Halimeda tuna, Codium bursa and Cystoseira barbata) and one seagrass (Cymodocea nodosa) were collected from the Coast of Montenegro, Gulf of Boka Kotorska and their chemical analysis was performed. In seagrass C. nodosa, three phenolic compounds were identified (diosmetin 7-sulfate, caftaric and coutaric acid). The content of ß-glucan, fatty acids, sterols and micro- and macro-elements were investigated among all samples. The highest content of ß-glucan was detected in C. nodosa seagrass (13.04±0.42 g/100 g). The highest polyunsaturated fatty acids (PUFAs) level was reported in C. barbata, the brown alga (7.157 mg/g), which also had the significant sterol content (fucosterol, 21.76±0.1 µg/g). Green algae, C. bursa and H. tuna, showed the highest level of sterols (ß-sitosterol, 95.21±0.16 µg/g and 73.90±0.08 µg/g, respectively). H. tuna had the highest content of calcium (Ca) in amount of 55125 µg/g. In C. bursa, C. barbata and C. nodosa, the Na/K ratio was low (0.43, 0.46 and 0.69, respectively).

Strategy of port baseline surveys (PBS) in the Adriatic Sea.

Port baseline surveys (PBS) provide species inventories in and around ports, with a focus on non-indigenous species that may have been introduced by vessels, primarily via ballast water. PBS are an essential tool to support effective management strategies for non-indigenous as well as native harmful aquatic organisms and pathogens (HAOP). This paper describes the methodology of PBS that were conducted in 12 Adriatic ports. The PBS employed existing protocols that were adapted to meet the characteristics of the Adriatic sites. Their results are reported in several papers included in this special issue, each of which is devoted to a specific community. An overview of existing surveys protocols - which provide valuable support to decision-making and to design effective monitoring of non-indigenous species - is also supplied.

3D chitinous scaffolds derived from cultivated marine demosponge Aplysina aerophoba for tissue engineering approaches based on human mesenchymal stromal cells.

The recently discovered chitin-based scaffolds derived from poriferans have the necessary prosperities for potential use in tissue engineering. Among the various demosponges of the Verongida order, Aplysina aerophoba is an attractive target for more in-depth investigations, as it is a renewable source of unique 3D microporous chitinous scaffolds. We found these chitinous scaffolds were cytocompatible and supported attachment, growth and proliferation of human mesenchymal stromal cells (hMSCs) in vitro. Cultivation of hMSCs on the scaffolds for 7days resulted in a two-fold increase in their metabolic activity, indicating increased cell numbers. Cells cultured onto chitin scaffolds in differentiation media were able to differentiate into the chondrogenic, adipogenic and osteogenic lineages, respectively. These results indicate A. aerophoba is a novel source of chitin scaffolds to futher hMSCs-based tissue engineering strategies.